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A re-evaluation of changes in rat striatal D-2 dopamine receptors during development and aging
Neurobiology of Aging, Vol. 7, pp. 265-267, 1986. ©Ankho InternationalInc. Printed in the U.S.A.
0197-4580/86 $3.00 + .00
A Re-Evaluation of Changes in Rat Striatal D-2 Dopamine Receptors During Development and Aging K A T H Y M. O'BOYLE A N D J O H N L . W A D D I N G T O N 1
Department of Clinical Pharmacology, Royal College o f Surgeons in Ireland St. Stephen's Green, Dublin 2, Ireland R e c e i v e d 26 N o v e m b e r 1985 O'BOYLE, K. M. AND J. L. WADDINGTON. A re-evaluation of changes in rat striatal 1)-2 dopamine receptors during development and aging. NEUROBIOL AGING 7(4) 265-267, 1986.--High affinity binding of 3H-spiperone to D-2 dopamine receptors was measured in membrane fractions prepared from striata of male Sprague-Dawley rats aged 0.75-25 months. 3H-Spiperone bound with approximately 0.11 nM affinity in all age groups studied. The maximum number.of D-2 receptors initially increased with age, peak adult levels being reached by 4 months of age. Receptor density declined by 33% over the next 7 months and no further loss of receptors was apparent between 11 and 25 months of age. Such a time-course suggests a loss of rat D-2 receptors that occurs, at least in this strain, earlier than has previously been appreciated, and appears to be in agreement with preliminary positron emission tomographic studies in living man. Aging
Development
3H-Spiperone binding
Senescence
Striatal D-2 dopamine receptors
tained from Biolabs (Ballina, Ireland). Six age-groups were studied: 0.75, 4, 6, 11, 22 and 25 months old.
A G E - R E L A T E D changes in striatal dopaminergic function have been proposed to contribute to the slowing of motor performance often observed in the elderly [5]. Several aspects of dopaminergic transmission deteriorate with advancing age. F o r example, the content of dopamine in dopaminergic neurones is decreased and the ability of the aged brain to synthesise dopamine is impaired [11]. Dopaminergic agents initiate their actions by attaching to specific cell membrane receptors of which there are two major classes, D-1 and D-2 [7]. Using radioligand binding techniques the various subtypes and affinity states of central dopamine receptors can be distinguished and many of these have been studied for possible age-related changes with some conflicting results. The most consistent finding about age-related changes in dopamine receptors has been that D-2 receptor density in the basal ganglia of several species including man is reduced [3, 8, 12, 14, 17]. To examine such changes in greater detail we have re-evaluated age-related alterations in dopaminergic D-2 receptors by examining the binding properties of 3H-spiperone in striatal homogenates derived from rats of various ages.
Drugs and Chemicals Tritiated ligand, (phenyl-4-3H)-spiperone, 15-16 Ci/mmol, was purchased from Amersham (U.K.). Domperidone was a gift from Janssen Pharmaceutica (Belgium).
Radioligand Binding Experiments Specific binding of 3H-spiperone (3H-SPIP) was measured as described previously [8] in striatal homogenates prepared from male Sprague-Dawley rats. Total binding of 3H-SPIP and non-specific binding in the presence of 1 /zM domperidone was determined in 50 mM Tris-HCl buffer (pH 7.6 at 25°C) containing 120 mM NaCI, 5 mM KCI, 1 mM MgCI2, 2 mM CaCI2, 0.2 mM Na~SzO5 and 10/xM pargyline. F o u r mg original wet weight of tissue was incubated with 0.05--1.90 nM 3H-SPIP, and domperidone when used, in a 5 ml incubation volume. After incubation at 37°C for 20 min tubes were filtered and washed through GF/B filters under reduced pressure. Individual saturation curves were constructed using 6 concentrations of 3H-SPIP, each in duplicate, on homogenates derived from single rats. Data were subjected to Scatchard analysis and Ka and Bmax values were calculated from best fit linear regressions. Data are presented as means_+ s.e.m.
METHOD
Animals Male Sprague-Dawley rats were obtained from the Wolfson Institute for Gerontology (Hull, U.K.), except for 0.75 month old animals of the same strain which were ob-
~Requests for reprints should be addressed to John L. Waddington.
265
266
O ' B O Y L E AND W A D D I N G T O N
(a)
(b) Kd
200
2O
1oOI+....+., 0~4)
O~
.......
(51 ('1'2)
, ................
,_..+
(3)
(5)
(|1)
e-
E e~
¢a
"•10
20 Bmax
e lOO
t~
15
mlu-
~J .m ¢J
+
+
1G
O. J
1-0
2-0
aH-spip nM
10
20
B pm/g
FIG. 1. (a) Saturation isotherms and (b) corresponding Scatchard plots of specific 3H-SPIP binding in striatal membranes prepared from infant (0.75 months, A), young adult (4 months, O) and senescent (25 months, ©) rats. Points are means of 4-11 independent experiments.
16 AGE
is
2'0
2'5
months
FIG. 2. Changes in rat striatal dopamine receptor characteristics (K~, nM; B. . . . pmol/g) with age (months). Points are means_+s.e.m. of (n) independent experiments.
Statistics Data were analysed by a one-way analysis of variance (ANOVA). Individual group differences were determined using the Newman-Keuls test. RESULTS
Effects of Age on D-2 Receptors 3H-SPIP binding in striata from animals aged 0.75, 4, 6, 11,22 and 25 months was studied. The wet weights of pooled right and left striata used in these binding assays did not change with age (e.g., 0.75 months: 0.093-+0.004 g, n=4; 25 months: 0.099-+0.006 g, n= 11), nor was there an age-related change in the protein to wet weight ratio in the two groups studied (4 months: 41.8-+2.8 mg protein/g wet weight, n = 11 ; 22 months: 37.8-+6.5 mg protein/g wet weight, n = l l ) . For each age group, binding was saturable and consistent with binding to a single population of sites. Figure 1 shows the saturation isotherms and corresponding scatchard plots for specific 3H-SPIP binding to infant (0.75 month), young adult (4 month) and senescent (25 month) striatal membranes. The overall relationship between age and the binding parameters of Kd and Bmax are shown in Fig. 2. A N O V A revealed that age had a significant effect on D-2 receptor number, F(5,34)=4.79, p<0.005. Between 0.75 and 4 months of age the number of D-2 receptors doubled, increasing from 8.73-+0.47 (n=4) to 17.6-+1.9 (n=5) pmol/g wet weight (p<0.05, Newman-Keuls). After 4 months of age receptor density began to decline, reaching a minimum level, corresponding to a 34% loss, at 11 months (Bma×=ll.6-+0.2 pmol/g, n=3, p < 0 . 0 5 vs. 4 month old rats). There was no further loss of receptors over the next 14 months of the rat's life (Bmax 25 months=ll.8-+0.9 pmol/g, n = l l ) . The Kd value did not change with age; in all experiments it ranged from 0.08 to 0.12 nM with an average of 0.11 nM (Fig. 2). DISCUSSION
The present study was undertaken to determine the pattern of change of D-2 dopamine receptors with age. To
achieve this, :+H-SPIP binding was measured in striatat homogenates derived from rats of 6 different ages. The particular ages of the rats studied included the major physiological landmarks for this species life-span. Thus, 0.75 month old rats can be described as "infant" [4]; the first 9 months can be described as " y o u t h " ; " m a t u r i t y " is reached after 10-12 months and '+senescence" begins to develop over the last quarter of life [11]. The aged animals used in this study have a maximum (rarely attained) life-span of approximately 28 months, 50% mortality occurring at 24 months. The results presented here indicate that there are two phases to the relationship between aging and D-2 receptors. Phase 1 occurs between infancy and early youth and during this time D-2 receptors appear to increase in number, reaching a value at 4 months which is twice the number present in 0.75 month old animals. Although the 0.75 and 4 month old male Sprague-Dawley rats were by necessity obtained from different sources, the present observation that D-2 receptors appear to increase in number during the first few months of life is in good agreement with the results of several previous studies in various strains [4, 6, 9]. While this study indicates that peak D-2 receptor density occurs by 4 months of age it is not possible to identify the exact age at which the true peak occurs without studying animals of intermediate ages. Postnatal development of dopamine receptors more closely parallels the appearance of cholinergic neurones and even of endogenous acetylcholine than it does the presynaptic markers for dopaminergic neurones [1] suggesting that these receptors are located post-synaptically on small cholinergic inter-neurons. However, development of these receptors is also linked to nerve terminal ontogeny since destruction of presynaptic dopaminergic terminals by 6-OHDA prevents the developmental increase in the number of dopamine receptor binding sites [2]. It is now reasonably well established that striatal D-2 receptor number is reduced in senescent rats [8,12]. However, many of these studies have confined their analyses to comparing young and aged animals. In the present study we have
DOPAMINE RECEPTORS DURING D E V E L O P M E N T AND A G I N G examined the age-related loss of D-2 receptors comparing animals aged 4, 6, 11, 22 and 25 months. Our results show that the density of D-2 receptors begins to decline after 4 months of age reaching a minimum level by II months of age. There was no further loss of receptors as the animals reached old age and senescence. These results in male Sprague-Dawley rats differ from those of others who reported a progressive reduction in receptor concentration over the life-span of the male Wistar rat [13]; also, there is a recent report that in male C57BL mice the high affinity form of the D-2 receptor declines in the first half of the life-span, while only the total D-2 receptor concentration continues to decline thereafter [15]. The existence of minor methodological differences (such as the use of domperidone instead of butaclamol to correct non-specific binding) between this and other studies make it impossible to state conclusively that the different patterns of receptor loss manifested by SpragueDawley and Wistar rats reflect a strain difference. Nevertheless, the apparent lack of dependence of mouse striatal D-2 receptor characteristics on the choice of domperidone vs. butaclamol as blank [15] supports the present interpretation of a strain difference. These results suggest a loss of striatal D-2 receptors in male Sprague-Dawley rats that occurs earlier than has previously been appreciated. This time scale of receptor loss may be similar in male Sprague-Dawley rats and male humans; measurement of D-2 receptors in the caudate/putamen of healthy males by in vivo positron emission tomography with J~C-N-methyl spiperone has suggested that the age-related decline in D-2 receptors may be almost complete by 45-50 years of age [17], though the age range of subjects is not uniform. In the same study, females showed a more linear decline in D-2 receptors with age and the authors suggest that there may be a sex difference in the loss of D-2 recep-
267
tors. Post-mortem studies on human D-2 receptors have yielded less clear cut results whereby D-2 sites decline progressively with age in the caudate nucleus but show no agerelated change in putamen tissue [14]. It is also noteworthy that the age-related loss of specific 3H-QNB binding to rat striatal muscarinic cholinergic receptors occurred between 3-6 months and 9-12 months of age [10]. This is interesting in view of the fact that these 3H-QNB labelled sites are located in the same brain region as the 3H-SPIP labelled D-2 sites. Reduction of dopamine binding sites in the basal ganglia could contribute to the age-related increase in movement disorders in humans [5]. There does, however, appear to be a temporal disparity between loss of receptors and appearance of motor disturbances; if, as reported here, D-2 receptors do indeed decline by midlife then this event may precede the appearance of motor disturbances by one quarter of the species life span. A possible explanation to account for this disparity may relate to the ability of the remaining receptors to compensate for any loss of total receptor number. In experiments designed to assess the compensatory capacity of rat striatum our results indicate that 25 month old rats have an impaired ability to develop receptor supersensitivity in response to 3 months chronic haloperidol treatment compared to their young counterparts [16]. This suggests that in this strain regulation of dopamine receptors in senescence may be compromised. Thus, mature rats that have fewer receptors than their young adult counterparts may not display motor deficits if their ability to respond to increased demand is intact. ACKNOWLEDGEMENTS This work was supported by the Medical Research Council of Ireland, the Royal College of Surgeons in Ireland, the Royal College of Physicians of Ireland, Sanity, Janssen and Squibb.
REFERENCES
1. Coyle, J. T. and P. Compochiaro. Ontogenesis of dopaminergic-cholinergic interactions in the rat striatum: a neurochemical study. J Neurochem 27: 673-678, 1976. 2. Deskin, R., F. J. Seidler, W. L. Whitmore and T. A. Slotkin. Development of a-noradrenergic and dopaminergic receptor systems depends on maturation of their presynaptic nerve terminals in the rat brain. J Neurochem 36: 1683-1690, 1981. 3. Finch, C. E., J. F. Marshall and P. K. Randall. Ageing and basal ganglia functions. Annu Rev Gerontol Geriatr 2: 4%87, 1981. 4. Hartley, E. J. and P. Seeman. Development of receptors for dopamine and noradrenaline in rat brain. Eur J Pharmacol 91: 391-397, 1983. 5. Hornykiewicz, O. Dopamine changes in the aging human brain: functional considerations. Aging 23: 9-14, 1983. 6. Karakiulakis, G., A. G. Paradelis, P. G. Papaioannidou and P. J. Thomas. Maturational aspects of the dopaminergic system: ontogenesis of high affinity dopamine binding to neural membrane fragments of the rat brain. Methods Find Exp Clin Pharmacol 5: 685-694, 1983. 7. Left, S. E. and I. Creese. Dopamine receptors reexplained. Trends Pharmacol Sci 4: 463--467, 1983. 8. O'Boyle, K. M. and J. L. Waddington. Loss of rat striatal dopamine receptors with ageing is selective for D-2 but not D-1 sites: association with increased non-specific binding of the D-1 ligand 3H-piflutixol. Eur J Pharmacol 105: 171-174, 1984. 9. Pardo, J. V., I. Creese, D. R. Burt and S. H. Snyder. Ontogenesis of dopamine receptor binding in the corpus striatum of the rat. Brain Res 125: 376-382, 1977. 10. Pedigo, N. W., Jr., L. D. Minor and T. N. Krumrei. Cholinergic drug effects and brain muscarinic receptor binding in aged rats. Neurobiol Aging 5: 227-233, 1984.
11. Pradhan, S. A. Central neurotransmitters and aging. Life Sci 26: 1643-1656, 1980. 12. Roth, G. S. and G. D. Hess. Changes in the mechanisms of hormone and neurotransmitter action during aging: current status of the role of receptor and post-receptor alterations. A review. Mech Ageing Dev 20: 175-194, 1982. 13. Roth, G. S., D. K. Ingram and J. A. Joseph. Delayed loss of striatal dopamine receptors during aging of dietarily restricted rats. Brain Res 300: 27-32, 1984. 14. Severson, J. A., J. Marcusson, B. Winblad and C. E. Finch. Age-correlated loss of dopaminergic binding sites in human basal ganglia. J Neurochem 39: 1623-1631, 1982. 15. Severson, J. A. and P. K. Randall. D-2 dopamine receptors in aging mouse striatum: determination of high- and low-affinity agonist binding sites. J Pharmacol Exp Ther 233: 361-368, 1985. 16. Waddington, J. L., K. M. O'Boyle, A. G. Molloy, H. A. Youssef, D. J. King and S. J. Cooper. Neurotransmitter receptors and ageing: dopamine/neuroleptic receptors, involuntary movements and the disease process of schizophrenia. In: Therapeutics in the Elderly: Scientific Foundations and Clinical Practice, edited by K. O'Malley and J. L. Waddington.
Amsterdam: Elsevier, 1985, pp. 63-76. 17. Wong, D. F., H. N. Wagner, Jr., R. F. Dannals, J. M. Links, J. J. Frost, H. T. Ravert, A. A. Wilson, A. E. Rosenbaum, A. Gjedde, K. H. Douglass, J. D. Petronis, M. F. Folstein, J. K. T. Toung, H. D. Burns and M. J. Kuhar. Effects of age on dopamine and serotonin receptors measured by positron tomography in the living human brain. Science 226: 1393-1396, 1984.
0197-4580/86 $3.00 + .00
A Re-Evaluation of Changes in Rat Striatal D-2 Dopamine Receptors During Development and Aging K A T H Y M. O'BOYLE A N D J O H N L . W A D D I N G T O N 1
Department of Clinical Pharmacology, Royal College o f Surgeons in Ireland St. Stephen's Green, Dublin 2, Ireland R e c e i v e d 26 N o v e m b e r 1985 O'BOYLE, K. M. AND J. L. WADDINGTON. A re-evaluation of changes in rat striatal 1)-2 dopamine receptors during development and aging. NEUROBIOL AGING 7(4) 265-267, 1986.--High affinity binding of 3H-spiperone to D-2 dopamine receptors was measured in membrane fractions prepared from striata of male Sprague-Dawley rats aged 0.75-25 months. 3H-Spiperone bound with approximately 0.11 nM affinity in all age groups studied. The maximum number.of D-2 receptors initially increased with age, peak adult levels being reached by 4 months of age. Receptor density declined by 33% over the next 7 months and no further loss of receptors was apparent between 11 and 25 months of age. Such a time-course suggests a loss of rat D-2 receptors that occurs, at least in this strain, earlier than has previously been appreciated, and appears to be in agreement with preliminary positron emission tomographic studies in living man. Aging
Development
3H-Spiperone binding
Senescence
Striatal D-2 dopamine receptors
tained from Biolabs (Ballina, Ireland). Six age-groups were studied: 0.75, 4, 6, 11, 22 and 25 months old.
A G E - R E L A T E D changes in striatal dopaminergic function have been proposed to contribute to the slowing of motor performance often observed in the elderly [5]. Several aspects of dopaminergic transmission deteriorate with advancing age. F o r example, the content of dopamine in dopaminergic neurones is decreased and the ability of the aged brain to synthesise dopamine is impaired [11]. Dopaminergic agents initiate their actions by attaching to specific cell membrane receptors of which there are two major classes, D-1 and D-2 [7]. Using radioligand binding techniques the various subtypes and affinity states of central dopamine receptors can be distinguished and many of these have been studied for possible age-related changes with some conflicting results. The most consistent finding about age-related changes in dopamine receptors has been that D-2 receptor density in the basal ganglia of several species including man is reduced [3, 8, 12, 14, 17]. To examine such changes in greater detail we have re-evaluated age-related alterations in dopaminergic D-2 receptors by examining the binding properties of 3H-spiperone in striatal homogenates derived from rats of various ages.
Drugs and Chemicals Tritiated ligand, (phenyl-4-3H)-spiperone, 15-16 Ci/mmol, was purchased from Amersham (U.K.). Domperidone was a gift from Janssen Pharmaceutica (Belgium).
Radioligand Binding Experiments Specific binding of 3H-spiperone (3H-SPIP) was measured as described previously [8] in striatal homogenates prepared from male Sprague-Dawley rats. Total binding of 3H-SPIP and non-specific binding in the presence of 1 /zM domperidone was determined in 50 mM Tris-HCl buffer (pH 7.6 at 25°C) containing 120 mM NaCI, 5 mM KCI, 1 mM MgCI2, 2 mM CaCI2, 0.2 mM Na~SzO5 and 10/xM pargyline. F o u r mg original wet weight of tissue was incubated with 0.05--1.90 nM 3H-SPIP, and domperidone when used, in a 5 ml incubation volume. After incubation at 37°C for 20 min tubes were filtered and washed through GF/B filters under reduced pressure. Individual saturation curves were constructed using 6 concentrations of 3H-SPIP, each in duplicate, on homogenates derived from single rats. Data were subjected to Scatchard analysis and Ka and Bmax values were calculated from best fit linear regressions. Data are presented as means_+ s.e.m.
METHOD
Animals Male Sprague-Dawley rats were obtained from the Wolfson Institute for Gerontology (Hull, U.K.), except for 0.75 month old animals of the same strain which were ob-
~Requests for reprints should be addressed to John L. Waddington.
265
266
O ' B O Y L E AND W A D D I N G T O N
(a)
(b) Kd
200
2O
1oOI+....+., 0~4)
O~
.......
(51 ('1'2)
, ................
,_..+
(3)
(5)
(|1)
e-
E e~
¢a
"•10
20 Bmax
e lOO
t~
15
mlu-
~J .m ¢J
+
+
1G
O. J
1-0
2-0
aH-spip nM
10
20
B pm/g
FIG. 1. (a) Saturation isotherms and (b) corresponding Scatchard plots of specific 3H-SPIP binding in striatal membranes prepared from infant (0.75 months, A), young adult (4 months, O) and senescent (25 months, ©) rats. Points are means of 4-11 independent experiments.
16 AGE
is
2'0
2'5
months
FIG. 2. Changes in rat striatal dopamine receptor characteristics (K~, nM; B. . . . pmol/g) with age (months). Points are means_+s.e.m. of (n) independent experiments.
Statistics Data were analysed by a one-way analysis of variance (ANOVA). Individual group differences were determined using the Newman-Keuls test. RESULTS
Effects of Age on D-2 Receptors 3H-SPIP binding in striata from animals aged 0.75, 4, 6, 11,22 and 25 months was studied. The wet weights of pooled right and left striata used in these binding assays did not change with age (e.g., 0.75 months: 0.093-+0.004 g, n=4; 25 months: 0.099-+0.006 g, n= 11), nor was there an age-related change in the protein to wet weight ratio in the two groups studied (4 months: 41.8-+2.8 mg protein/g wet weight, n = 11 ; 22 months: 37.8-+6.5 mg protein/g wet weight, n = l l ) . For each age group, binding was saturable and consistent with binding to a single population of sites. Figure 1 shows the saturation isotherms and corresponding scatchard plots for specific 3H-SPIP binding to infant (0.75 month), young adult (4 month) and senescent (25 month) striatal membranes. The overall relationship between age and the binding parameters of Kd and Bmax are shown in Fig. 2. A N O V A revealed that age had a significant effect on D-2 receptor number, F(5,34)=4.79, p<0.005. Between 0.75 and 4 months of age the number of D-2 receptors doubled, increasing from 8.73-+0.47 (n=4) to 17.6-+1.9 (n=5) pmol/g wet weight (p<0.05, Newman-Keuls). After 4 months of age receptor density began to decline, reaching a minimum level, corresponding to a 34% loss, at 11 months (Bma×=ll.6-+0.2 pmol/g, n=3, p < 0 . 0 5 vs. 4 month old rats). There was no further loss of receptors over the next 14 months of the rat's life (Bmax 25 months=ll.8-+0.9 pmol/g, n = l l ) . The Kd value did not change with age; in all experiments it ranged from 0.08 to 0.12 nM with an average of 0.11 nM (Fig. 2). DISCUSSION
The present study was undertaken to determine the pattern of change of D-2 dopamine receptors with age. To
achieve this, :+H-SPIP binding was measured in striatat homogenates derived from rats of 6 different ages. The particular ages of the rats studied included the major physiological landmarks for this species life-span. Thus, 0.75 month old rats can be described as "infant" [4]; the first 9 months can be described as " y o u t h " ; " m a t u r i t y " is reached after 10-12 months and '+senescence" begins to develop over the last quarter of life [11]. The aged animals used in this study have a maximum (rarely attained) life-span of approximately 28 months, 50% mortality occurring at 24 months. The results presented here indicate that there are two phases to the relationship between aging and D-2 receptors. Phase 1 occurs between infancy and early youth and during this time D-2 receptors appear to increase in number, reaching a value at 4 months which is twice the number present in 0.75 month old animals. Although the 0.75 and 4 month old male Sprague-Dawley rats were by necessity obtained from different sources, the present observation that D-2 receptors appear to increase in number during the first few months of life is in good agreement with the results of several previous studies in various strains [4, 6, 9]. While this study indicates that peak D-2 receptor density occurs by 4 months of age it is not possible to identify the exact age at which the true peak occurs without studying animals of intermediate ages. Postnatal development of dopamine receptors more closely parallels the appearance of cholinergic neurones and even of endogenous acetylcholine than it does the presynaptic markers for dopaminergic neurones [1] suggesting that these receptors are located post-synaptically on small cholinergic inter-neurons. However, development of these receptors is also linked to nerve terminal ontogeny since destruction of presynaptic dopaminergic terminals by 6-OHDA prevents the developmental increase in the number of dopamine receptor binding sites [2]. It is now reasonably well established that striatal D-2 receptor number is reduced in senescent rats [8,12]. However, many of these studies have confined their analyses to comparing young and aged animals. In the present study we have
DOPAMINE RECEPTORS DURING D E V E L O P M E N T AND A G I N G examined the age-related loss of D-2 receptors comparing animals aged 4, 6, 11, 22 and 25 months. Our results show that the density of D-2 receptors begins to decline after 4 months of age reaching a minimum level by II months of age. There was no further loss of receptors as the animals reached old age and senescence. These results in male Sprague-Dawley rats differ from those of others who reported a progressive reduction in receptor concentration over the life-span of the male Wistar rat [13]; also, there is a recent report that in male C57BL mice the high affinity form of the D-2 receptor declines in the first half of the life-span, while only the total D-2 receptor concentration continues to decline thereafter [15]. The existence of minor methodological differences (such as the use of domperidone instead of butaclamol to correct non-specific binding) between this and other studies make it impossible to state conclusively that the different patterns of receptor loss manifested by SpragueDawley and Wistar rats reflect a strain difference. Nevertheless, the apparent lack of dependence of mouse striatal D-2 receptor characteristics on the choice of domperidone vs. butaclamol as blank [15] supports the present interpretation of a strain difference. These results suggest a loss of striatal D-2 receptors in male Sprague-Dawley rats that occurs earlier than has previously been appreciated. This time scale of receptor loss may be similar in male Sprague-Dawley rats and male humans; measurement of D-2 receptors in the caudate/putamen of healthy males by in vivo positron emission tomography with J~C-N-methyl spiperone has suggested that the age-related decline in D-2 receptors may be almost complete by 45-50 years of age [17], though the age range of subjects is not uniform. In the same study, females showed a more linear decline in D-2 receptors with age and the authors suggest that there may be a sex difference in the loss of D-2 recep-
267
tors. Post-mortem studies on human D-2 receptors have yielded less clear cut results whereby D-2 sites decline progressively with age in the caudate nucleus but show no agerelated change in putamen tissue [14]. It is also noteworthy that the age-related loss of specific 3H-QNB binding to rat striatal muscarinic cholinergic receptors occurred between 3-6 months and 9-12 months of age [10]. This is interesting in view of the fact that these 3H-QNB labelled sites are located in the same brain region as the 3H-SPIP labelled D-2 sites. Reduction of dopamine binding sites in the basal ganglia could contribute to the age-related increase in movement disorders in humans [5]. There does, however, appear to be a temporal disparity between loss of receptors and appearance of motor disturbances; if, as reported here, D-2 receptors do indeed decline by midlife then this event may precede the appearance of motor disturbances by one quarter of the species life span. A possible explanation to account for this disparity may relate to the ability of the remaining receptors to compensate for any loss of total receptor number. In experiments designed to assess the compensatory capacity of rat striatum our results indicate that 25 month old rats have an impaired ability to develop receptor supersensitivity in response to 3 months chronic haloperidol treatment compared to their young counterparts [16]. This suggests that in this strain regulation of dopamine receptors in senescence may be compromised. Thus, mature rats that have fewer receptors than their young adult counterparts may not display motor deficits if their ability to respond to increased demand is intact. ACKNOWLEDGEMENTS This work was supported by the Medical Research Council of Ireland, the Royal College of Surgeons in Ireland, the Royal College of Physicians of Ireland, Sanity, Janssen and Squibb.
REFERENCES
1. Coyle, J. T. and P. Compochiaro. Ontogenesis of dopaminergic-cholinergic interactions in the rat striatum: a neurochemical study. J Neurochem 27: 673-678, 1976. 2. Deskin, R., F. J. Seidler, W. L. Whitmore and T. A. Slotkin. Development of a-noradrenergic and dopaminergic receptor systems depends on maturation of their presynaptic nerve terminals in the rat brain. J Neurochem 36: 1683-1690, 1981. 3. Finch, C. E., J. F. Marshall and P. K. Randall. Ageing and basal ganglia functions. Annu Rev Gerontol Geriatr 2: 4%87, 1981. 4. Hartley, E. J. and P. Seeman. Development of receptors for dopamine and noradrenaline in rat brain. Eur J Pharmacol 91: 391-397, 1983. 5. Hornykiewicz, O. Dopamine changes in the aging human brain: functional considerations. Aging 23: 9-14, 1983. 6. Karakiulakis, G., A. G. Paradelis, P. G. Papaioannidou and P. J. Thomas. Maturational aspects of the dopaminergic system: ontogenesis of high affinity dopamine binding to neural membrane fragments of the rat brain. Methods Find Exp Clin Pharmacol 5: 685-694, 1983. 7. Left, S. E. and I. Creese. Dopamine receptors reexplained. Trends Pharmacol Sci 4: 463--467, 1983. 8. O'Boyle, K. M. and J. L. Waddington. Loss of rat striatal dopamine receptors with ageing is selective for D-2 but not D-1 sites: association with increased non-specific binding of the D-1 ligand 3H-piflutixol. Eur J Pharmacol 105: 171-174, 1984. 9. Pardo, J. V., I. Creese, D. R. Burt and S. H. Snyder. Ontogenesis of dopamine receptor binding in the corpus striatum of the rat. Brain Res 125: 376-382, 1977. 10. Pedigo, N. W., Jr., L. D. Minor and T. N. Krumrei. Cholinergic drug effects and brain muscarinic receptor binding in aged rats. Neurobiol Aging 5: 227-233, 1984.
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